Clostridioides difficile is a gram-positive, spore forming, anaerobic bacteria that is the cause of the most common healthcare associated infection. C. difficile infections (CDI) are estimated to cause nearly 450,000 infections and 30,000 deaths within the United States per year. Since the early 2000s, the epidemic strain identified as restriction endonuclease analysis (REA) group BI, PCR ribotype (RT) 027 (BI/RT027) has been the most common cause of CDI in the United States. However, in the past 10 years there has been a drastic change in the molecular epidemiology of C. difficile as BI/RT027 prevalence has decreased. Furthermore, there is evidence that healthcare-associated CDI (HA-CDI) is on the decline and community-associated CDI (CA-CDI) is increasing. One strain identified as REA group Y, which commonly correlates to RT 014 and 020 [Y(RT014/020)], has been endemic in the United States accounting for >5% of all CDI since the 1980s. As BI/RT027 associated CDI has decreased over the past 10 years, Y(RT014/020) related CDI have steadily increased and now account for 10-15% of all CDI. With the rise in CA-CDI, there is rising evidence that Y(RT014/020) resides within an unknown community reservoir. Despite a decades long prevalence and increasing concern with CA-CDI, a detailed molecular analysis of Y(RT014/020) has never been completed. Therefore, our proposal will be to complete a detailed molecular and clinical analysis in conjunction with a rigorous genomic analysis of Y(RT014/020) and BI/RT027 to assess the factors which contribute towards our overarching hypothesis that Y(RT014/020) resides within an unknown community reservoir. Support from the career development award (CDA) will provide Dr. Skinner with the necessary foundation and training to establish himself as a successful VA biomedical and laboratory clinician-scientist focused on the pathogenesis and epidemiology of C. difficile. The molecular epidemiology of C. difficile has continually shifted over multiple decades. These shifts are the results of underlying pathologic factors which influence strain selection. This innovative project will utilize C. difficile from multiple decades to understand the factors which have allowed for one strain (REA group Y) to remain endemic for decades. This information could be further utilized to understand which factors (i.e., antibiotic exposure) place Veterans at risk for developing a CDI and provide a deeper understanding into the mechanisms by which C. difficile is resistant to particular antibiotics. Therefore, the completion of this CDA will provide a critical foundation for Dr. Skinner and results from this project will be used to support Dr. Skinner's VA Merit Review application to study the mechanism by which C. difficile is resistant to certain cephalosporin antimicrobials.